Project Summary Cartilage tissue engineering techniques has advanced towards clinical applications. Our research has shown that engineered cartilage plugs can be cultivated in the laboratory with structural and functional properties towards healthy native tissue. Translating these findings to cultivating de novo tissues with human cells has been a significant challenge for the field; however, due to donor age and disease state of the tissue used to acquire cells. Thus, the overarching goal of this project is to use a novel high-throughput approach for determining tissue-growth potential of cell populations for cartilage tissue engineering and regeneration. We will determine elastic and viscoelastic cell mechanics using a low-cost microfluidic device designed in Dr. Lydia Sohn?s laboratory (co-I on this proposal). In contrast to other methods for determining cell mechanics, such as atomic force microscopy, our approach is not limited to small sample sizes (n < 50), does not require additional expensive equipment, and provides a fast assessment of cell mechanics (order of minutes) prior to cell-based tissue engineering culture without damage to the cells. In the first aim, we will determine elastic and viscoelastic cell mechanics using high-throughput screening and real-time analysis. We will optimize a high-throughput mechano-phenotyping device developed in Dr. Sohn?s laboratory to evaluate cell mechanics for cartilage tissue engineering. In the second aim, we will evaluate the relationship between cell mechanics and 3D tissue growth in engineered cartilage. We will evaluate tissue growth from primed and unprimed chondrocytes and MSCs encapsulated in agarose. Tissue culture studies will be performed with cells at Passage 2 and 4. The results from Aim 1 will guide our expected results for Aim 2; however, both passages (P2 and P4) and primed conditions (unprimed and primed) will be evaluated in 3D culture to confirm our hypothesis for Aim 2. Successful completion of this proposal has potential to have a transformative impact on cartilage tissue engineering outcomes. That is, a low-cost high-throughput device capable of measuring elastic and viscoelastic cell mechanics, could 1) decrease time between cell harvest and unknown 3D matrix production and 2) can be used as a future screening method for methods that aim to differentiate other cell sources towards chondrogenesis.